VACCINE. Development of a novel yeast-based oral subunit vaccine against Eimeria spp. in chickens

酵母种类，如酿酒酵母菌是高效生产重组蛋白的有用工具，具有公认的安全（GRAS）地位。它们能够进行许多其他表达系统无法实现的复杂的翻译后修饰，并且很容易生长到非常高的密度，产生大量稳定的颗粒。最近，利用酿酒葡萄球菌作为癌症、病毒和细菌疫苗的递送载体的想法已经被探索，诱导强大的体液和细胞免疫反应。除了使用酵母生产感兴趣的疫苗抗原外，酵母细胞本身已被证明具有佐剂样特性，并具有激活抗原呈递细胞上表达的炎症和吞噬受体的潜力。我们的初步数据表明，冷冻干燥的重组酵母培养物在其表面表达病毒蛋白，使重组酵母完全无法存活（未发表）。然而，冷冻干燥过程不会改变这些蛋白质的构象，因为通过流式细胞术和Western blotting量化，活酵母和冻干酵母的表面表达是相同的。这对疫苗设计具有有趣的意义，因为不能存活的酿酒葡萄球菌没有被归类为转基因生物（GMO），而且这种灭活疫苗不受转基因生物法规的约束。此外，冻干酵母不需要冷藏，从而降低了运输和储存成本。以酿酒葡萄球菌为基础的疫苗可能特别有价值，用于预防养殖家禽的疾病，其中安全性、可扩展性、稳定性、交付和成本至关重要。例如，现代家禽生产依赖于对艾美耳球虫的有效控制，但目前使用药物和活疫苗的方法需要改进。近年来，一组球虫抗原已被确定为候选疫苗，每一种抗原都能使卵囊产量减少65%。现在，重点正在从抗原发现转移到抗原配方和递送，酵母是一个主要的选择，特别是考虑到t细胞介导的反应在抗球虫免疫中的重要性。

Yeast species such as Saccharomyces cerevisiae are useful tools for the high yield production of recombinant proteins and have a Generally Regarded As Safe (GRAS) status. They are capable of performing several complex post-translational modifications that are not achieved in many other expression systems, and are easily grown to very high densities producing large quantities of stable particles. Recently, the idea of using S. cerevisiae as a delivery vehicle for cancer, viral, and bacterial vaccines has been explored, inducing robust humoral and cellular immune responses. In addition to using yeast to produce a vaccine antigen of interest, the yeast cell itself has been shown to have adjuvant-like properties and has the potential to activate both inflammatory and phagocytic receptors expressed on antigen-presenting cells. Our preliminary data demonstrates that freeze-drying recombinant yeast cultures expressing viral protein at their surface renders the recombinant yeast completely non-viable (unpublished). However, the freeze-drying process does not alter conformation of these proteins, as surface expression is equal in live and freeze-dried yeast as quantified by flow cytometry and Western blotting. This has interesting implications in vaccine design as a non-viable S. cerevisiae is not categorised as a genetically modified organism (GMO) and such a killed vaccine would not be subject to GMO regulations. Additionally there would be no need for refrigeration of the freeze-dried yeast, reducing transport and storage costs. Vaccines based upon S. cerevisiae are likely to be particularly valuable against diseases of farmed poultry, where safety, scalability, stability, delivery and cost are crucial. In one example modern poultry production relies on effective control of Eimeria, but current approaches using drugs and live vaccines require improvement. In recent years a panel of coccidial antigens have been identified as vaccine candidates, each individually capable of inducing up to a 65% reduction in oocyst output. Now, the focus is shifting from antigen discovery to antigen formulation and delivery with yeast being a leading option, especially given the importance of T-cell mediated responses in anticoccidial immunity.